Extended Data Fig. 10 | Validation of the PLXND1 mutant. a, Negative-stain
two-dimensional class averages of wild-type PLXND1 were obtained by
classifying 1,305 particles into 10 classes. Scale bar, 10 nm. b, Negative-stain
two-dimensional class averages of mutant PLXND1 were obtained by
classifying 1,357 particles into 10 classes. c, The double mutant of PLXND1 was
labelled with a thiol-reactive f luorescent dye, Alexa Fluor 488 C5 maleimide.
The degree of labelling shows that the vast majority of PLXND1 mutant
molecules form the disulfide-linked bond and thus the ring of the majority of
PLXND1-mutant molecules appears to be locked by the covalent bond. The
degree of labelling for the hen egg ovalbumin, which we used as a positive
control, is close to the number of free cysteines in ovalbumin. n = 3
independent experiments. Data are mean ± s.e.m. P values were calculated by
two-tailed Student’s t-tests using GraphPad Prism. ***P < 0.001.